The antioxidant activity of South African wines in different test systems as affected by cultivar and

252 Pages·2003·2.31 MB·English

by  De Beer

THE ANTIOXIDANT ACTIVITY OF SOUTH
AFRICAN WINES IN DIFFERENT TEST
SYSTEMS AS AFFECTED BY CULTIVAR
AND AGEING

DALENE DE BEER

Thesis presented in partial fulfilment of the requirements for the degree of

MASTER OF SCIENCE IN FOOD SCIENCE

in the Department of Food Science,
Faculty of Agricultural and Forestry Sciences,
University of Stellenbosch

Study Leader: Dr E. Joubert
Co-study Leaders: Dr W.C.A. Gelderblom
Dr M. Manley

March 2002
ii
DECLARATION

I, the undersigned hereby declare that the work contained in this thesis is my
own original work and has not been previously in its entirety or in part
submitted at any university for a degree.

_______________________ ___________________

Dalene de Beer Date
iii
ABSTRACT

Phenolic compounds in wine, due to their antioxidant activity, are reportedly
responsible for the health-promoting properties of wines. The effect of cultivar
and in-bottle ageing on the antioxidant activity of South African wines in
different types of antioxidant assays was, therefore, investigated.
The antioxidant activity of commercial South African red (Cabernet
Sauvignon, Ruby Cabernet, Pinotage, Shiraz, Merlot) and white (Sauvignon
blanc, Chenin blanc, Chardonnay, Colombard) cultivar wines was compared
using the 2,2’-azino-di-(3-ethylbenzothialozine-sulphonic acid) radical cation
(ABTS•+) scavenging, 2,2-diphenyl-1-picrylhydrazyl radical (DPPH•)
scavenging and microsomal lipid peroxidation (MLP) assays. The red wines
was more effective than the white wines on an “as-is” and an equal total
phenol content. The total antioxidant activity (TAA and TAA ) of Ruby
ABTS DPPH
Cabernet was the lowest of the red wines, but the antioxidant potency (AP
ABTS
and AP ) of red wine phenolic fractions did not differ (P ‡ 0.05). Ruby
DPPH
Cabernet and Pinotage were the least effective inhibitors of MLP, while Merlot
was the most effective of the red wines. Pinotage phenolic fractions had
lower (P < 0.05) AP than that of other red wines. Of the white wines,
MLP
Chardonnay and Chenin blanc had the highest and lowest effectivity
respectively according to all antioxidant parameters. Ascorbic acid present in
some wines increased and decreased their TAA and % MLP inhibition
respectively. TAA and % MLP inhibition correlated well (r ‡ 0.7, P < 0.001)
with total phenol content of red and white wines, as well as with flavanol
content of red wines and tartaric acid ester content of white wines. The
% MLP inhibition also correlated well with flavanol content of white wines. No
correlation (P > 0.01) was obtained between TAA or % MLP inhibition and
monomeric anthocyanin content of red wines. In the deoxyribose assay, red
wines were more pro-oxidant and exhibited lower hydroxyl radical scavenging
and metal chelating abilities than white wines.
The effect of in-bottle ageing on antioxidant activity of wines was
determined using the ABTS•+ and DPPH• scavenging assays. The TAA and
total phenol content of experimental red (Pinotage and Cabernet Sauvignon)
iv
and white (Chardonnay and Chenin blanc) cultivar wines, decreased
(P < 0.05) during 12 months of storage at 0, 15 and 30 ºC. The TAA of
ABTS
Cabernet Sauvignon and Chardonnay, stored at 30 ºC were lower (P < 0.05)
than at 0 ºC. The AP and AP of most wines also decreased during
ABTS DPPH
storage. The monomeric anthocyanin content of red wines decreased
(P < 0.05) rapidly at 15 and 30 ºC. The flavanol content of wines (except
Chenin blanc) increased during the first 9 months, decreasing again after 12
months, while minor changes in the flavonol and tartaric acid ester content of
both red and white wines were observed. The TAA exhibited a good
ABTS
correlation (r ‡ 0.7, P < 0.001) with total phenol content of red and white
wines, as well as with flavonol and tartaric acid ester content of red and white
wines and flavanol content of white wines. The monomeric anthocyanin
content of red wines correlated (r = 0.50, P < 0.001) weakly with TAA .
ABTS
The decrease in the TAA of wines could thus be mainly attributed to a
ABTS
decrease in their total phenol content.
v
UITTREKSEL

Die antioksidant aktiwiteit van fenoliese komponente in wyn is waarskynlik
verantwoordelik vir die gesondheidsvoordele daarvan. Die studie het dus
gepoog om effek van kultivar en veroudering na bottelering op die
antioksidant aktiwiteit van Suid-Afrikaanse wyne te ondersoek.
Die antioksidant aktiwiteit van kommersiële Suid-Afrikaanse rooi
(Cabernet Sauvignon, Ruby Cabernet, Pinotage, Shiraz, Merlot) en wit
(Sauvignon blanc, Chenin blanc, Chardonnay, Colombard) kultivarwyne is
vergelyk deur middel van die 2,2’-azino-di-(3-etielbensotialosien-sulfoon
suur)-radikaal katioon (ABTS•+) vernietigingstoets, 2,2-difeniel-1-pikriel-
•
hidrasielradikaal (DPPH ) vernietigingstoets en mikrosomale lipied-
peroksidasietoets (MLP). Die antioksidant aktiwiteit en die antioksidant
kragtigheid (AK) van die rooiwyne was beter as dié van witwyne in al drie
antioksidant toetse. Die totale antioksidant aktiwiteit (TAA en TAA )
ABTS DPPH
van Ruby Cabernet was die laagste van die rooiwyne, terwyl die AK en
ABTS
AK van rooiwyn fenoliese fraksies nie van mekaar verskil (P ‡ 0.05) het
DPPH
nie. Van die rooiwyne, het Ruby Cabernet en Pinotage die laagste en Merlot
die hoogste effektiwiteit in die MLP toets getoon. Die AK van Pinotage se
MLP
fenoliese fraksies was die laagste van die rooiwyne. Die witwyne,
Chardonnay en Chenin blanc, het onderskeidelik die beste en swakste
antioksidant aktiwiteit en AK van die witwyne getoon in al drie antioksidant
toetse. Askorbiensuur wat in sommige witwyne voorgekom het, het die TAA
van hierdie wyne verhoog, maar hul % MLP inhibisie verlaag. Die TAA en %
MLP inhibisie het goed gekorreleer (r ‡ 0.7, P < 0.001) met die totale
fenolinhoud van rooi- en witwyne, asook die flavanolinhoud van rooiwyne en
die wynsteensuur-esterinhoud van witwyne. Die % MLP inhibisie het ook
goed gekorreleer met die flavanolinhoud van witwyne. Geen korrelasie
(P > 0.1) is waargeneem tussen antioksidant aktiwiteit van rooiwyne en hul
monomeriese antosianien-inhoud. Rooiwyn was meer pro-oksidatief in die
deoksieribose toets as witwyne, maar was die swakste hidroksieradikaal-
vernietigers en metaalcheleerders.
vi
Die effek van veroudering na bottelering op die antioksidant aktiwiteit
van wyne soos bepaal met die ABTS•+ en DPPH• vernietigingstoetse, is
ondersoek. Die TAA en die totale fenolinhoud van eksperimentele rooi-
(Pinotage en Cabernet Sauvignon) en witwyne (Chardonnay en Chenin blanc)
het afgeneem (P < 0.05) tydens opberging na bottelering by 0, 15 en 30 ºC
oor 12 maande. Opberging by 30 ºC het ‘n groter vermindering (P < 0.05) in
die TAA waarde vir Cabernet Sauvignon en Chardonnay veroorsaak as by
ABTS
0 ºC. Die meeste wyne se AP en AP waardes het ook verminder
ABTS DPPH
(P < 0.05) na 12 maande. Drastiese vermindering (P < 0.05) in die
monomeriese antosianieninhoud van rooiwyne is opgemerk tydens opberging
by 15 en 30 ºC. Tydens die eerste 9 maande se opberging het die
flavanolinhoud van wyne toegeneem (P < 0.05) en daarna afgeneem
(P < 0.05) tot by 12 maande, terwyl flavonol- en wynsteensuuresterinhoud van
beide rooi- en witwyne min verandering ondergaan het. Die totale fenolinhoud
van rooi- en witwyne, asook die flavonol en wynsteensuur-esterinhoud van
rooi-en witwyne en die flavanolinhoud van witwyne, het goed gekorreleer
(r ‡ 0.7, P < 0.001) met die TAA . In teenstelling met die resultate vir
ABTS
kommersiële kultivarwyne, was die TAA van rooiwyne swak gekorreleer
ABTS
(r = 0.5, P < 0.001) met hul monomeriese antosianieninhoud. Die afname in
TAA van wyne tydens veroudering kon dus meestal toegeskryf word aan
ABTS
die afname in hul totale fenolinhoud.
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CONTENTS

Chapter Page

Abstract iii

Uittreksel v

List of abbreviations xii

Acknowledgements xiv

1 INTRODUCTION 1

2 PHENOLIC COMPOUNDS: A REVIEW OF THEIR
POSSIBLE ROLE AS IN VIVO ANTIOXIDANT
COMPONENTS OF WINE 9
2.1 Introduction 9
2.2 Reactive oxygen species and free radicals 10
2.3 Lipid peroxidation 12
2.3.1 Free radical chain reaction mechanism 12
2.3.2 Reaction kinetics 13
2.3.3 Initiation of lipid peroxidation 17
2.3.4 Products of lipid peroxidation 19
2.4 Antioxidants 19
2.4.1 Phenolic acids 20
2.4.2 Flavonoids 20
2.4.2.1 Flavanols 20
2.4.2.2 Flavonols 22
2.4.2.3 Anthocyanidins 22
2.4.2.4 Proanthocyanidins 24
2.5 Measurement of in vitro antioxidant activity 24
2.5.1 Radical scavenging test systems 24
viii
2.5.1.1 DPPH radical scavenging assay 27
2.5.1.2 ABTS radical cation scavenging assay 29
2.5.2 Biologically relevant test systems 30
2.5.2.1 Thiobarbituric acid reaction 31
2.5.2.2 Low-density lipoprotein peroxidation 32
2.5.2.3 Microsomal lipid peroxidation 35
2.5.2.4 Deoxyribose assay 37
2.6 Structure-activity relationships 39
2.6.1 General considerations 39
2.6.2 Phenolic acids 42
2.6.3 Flavonoids 44
2.6.4 Comparative activities in selected antioxidant assays 48
2.7 Absorption and bioavailability 50
2.8 Methods for estimation of phenolic composition 63
2.8.1 Total phenols 63
2.8.2 Anthocyanins 64
2.8.3 Flavanols 64
2.8.4 Flavonols and tartaric acid esters 65
2.9 Phenolic composition of wine 65
2.9.1 Effect of phenolic composition of grapes 68
2.9.2 Effect of vinification processes 69
2.9.3 Effect of maturation 70
2.9.3.1 Maturation in wooden barrels 70
2.9.3.2 In-bottle ageing 71
2.10 Summary 72
2.11 References 72

3 ANTIOXIDANT ACTIVITY OF SOUTH AFRICAN RED
AND WHITE CULTIVAR WINES: FREE RADICAL
SCAVENGING ACTIVITY 93
3.1 Introduction 93
3.2 Materials and methods 95
3.2.1 Wines and chemicals 95
3.2.2 Sample preparation 98
ix
3.2.3 Determining the phenolic composition 98
3.2.4 Measurement of the 2,2’-azino-di-(3-ethylbenzo-
thialozine-sulphonic acid) radical cation (ABTS•+)
scavenging activity 99
3.2.5 Measurement of the 2,2-diphenyl-1-picrylhydrazyl radical
•
(DPPH ) scavenging activity 102
3.2.6 Determination of the contribution of ascorbic acid to the
free radical scavenging activity 104
3.2.7 Statistical analysis 104
3.3 Results and discussion 104
3.3.1 Phenolic composition 104
3.3.2 Total antioxidant activity measured using the ABTS•+
scavenging assay 109
•
3.3.3 Antioxidant activity measured using the DPPH
scavenging assay 114
3.3.4 Contribution of ascorbic acid to the free radical
scavenging activity 116
3.3.5 Correlation analysis 118
3.3.6 Canonical discriminant analysis 123
3.4 Conclusions 126
3.5 References 127

4 ACTIVITY OF SOUTH AFRICAN RED AND WHITE
CULTIVAR WINES AND SELECTED WINE PHENOLIC
COMPOUNDS IN A MICROSOMAL LIPID
PEROXIDATION AND DEOXYRIBOSE ASSAY 133
4.1 Introduction 134
4.2 Materials and methods 135
4.2.1 Wines and chemicals 135
4.2.2 Sample preparation 136
4.2.3 Determining the phenolic composition and ascorbic acid
content 136
4.2.4 Microsomal lipid peroxidation (MLP) assay 136
x
4.2.5 Deoxyribose assay 139
4.2.6 Statistical analysis 141
4.3 Results and discussion 141
4.3.1 Phenolic composition and ascorbic acid content 141
4.3.2 Inhibition of microsomal lipid peroxidation (MLP) 142
4.3.3 Correlation analysis 145
4.3.4 Determination of IC values for selected wine phenolic
50
compounds 152
4.3.5 Activity in the deoxyribose assay 155
4.4 Conclusions 162
4.5 References 163

5 THE EFFECT OF IN-BOTTLE AGEING ON THE
ANTIOXIDANT ACTIVITY OF RED AND WHITE WINES 169
5.1 Introduction 170
5.2 Materials and methods 172
5.2.1 Wines and chemicals 172
5.2.2 Determining the phenolic composition 172
5.2.3 Measurement of the 2,2’-azino-di-(3-ethylbenzo-
thialozine-sulphonic acid) radical cation (ABTS•+) and
•
2,2-diphenyl-1-picrylhydrazyl radical (DPPH ) scavenging
activity 173
5.2.4 Statistical analysis 173
5.3 Results and discussion 173
5.3.1 Phenolic composition 173
5.3.1.1 Total phenol content 173
5.3.1.2 Anthocyanin content 177
5.3.1.3 Flavanol content 182
5.3.1.4 Flavonol content 185
5.3.1.5 Tartaric acid ester content 186
5.3.2 Total antioxidant activity measured using the ABTS•+ and
•
DPPH scavenging assay 187
5.3.3 Correlation analysis 193